Follicle-stimulating hormone (henceforth also abbreviated as “FSH”) is a hormone synthesized and secreted by gonadotrophic hormone-producing cells of the anterior pituitary. FSH has an action of stimulating growth of immature ovarian follicles and ripening the cells in the ovary. As a medicament, FSH has been used as an ovulation-inducing agent in the assisted reproductive technology (ART).
Preparations of human menopausal gonadotropin (hMG) extracted from postmenopausal women's urine have been conventionally used as FSH-containing compositions. The hMG preparations contain FSH and lutenizing hormone (LII) in which the activity ratio of FSH and LH is about 1:1. In foreign countries, “recombinant FSH preparations” produced by gene recombination without using human urine as a raw material have recently been mainly used, and also in Japan, Follistim (registered trademark) was approved in 2005. Since these recombinant preparations do not use urine as a raw material, they are featured not to contain impurities and have consistent quality.
Bio-active proteins are generally unstable in a state of an aqueous solution, and this tendency is enhanced when purity of a protein becomes higher. In aqueous solutions, proteins are hydrolyzed in the same manner as low molecular weight compounds, as well as cause chemical changes such as racemization, and further cause changes of higher-order structures (physical changes) since they are polymer compounds. Examples of such physical changes include denaturation, aggregation, adsorption, precipitation, and the like. In the process of the physical change, it is considered that denaturation occurs as a trigger, and such phenomena as aggregation, adsorption and precipitation are subsequently caused. The denaturation means changes of the three-dimensional structures (tertiary and quaternary structures) such as unfolding of folded chains, and since the denaturation leads loss of physiological activity in most cases, it is important to prevent denaturation for enhanced stability of protein or peptide preparations.
In order to suppress chemical and physical changes in protein preparations, contaminant proteins such as human serum albumin have been generally added as a stabilizing agent. However, to avoid the risk of contamination of viruses and the like, use of recombinant preparations has recently become the mainstream. For the recombinant preparations, various stabilization methods have been examined as substitutes for the addition of contaminant proteins.
For example, it has been investigated to increase stability of protein preparations by adding a compound having an action of forming a hydrogen bond with a protein molecule or enhancing hydration of a protein molecule, such as saccharides, surfactants, and amino acids (see, for example, Bull. Chem. Soc. Jpn., 53, pp. 2451-2455, 1980; J. Biol. Chem., 256, pp. 7193-7201, 1981; Pharm. Res., 10, pp. 954-962, 1993; Int. J. Pharm., 96, pp. 41-49, 1993; Pharm. Res., 8, pp. 1258-1263, 1991; Pharm. Res., 10, pp. 649-659, 1993, and the like). As for amino acids among such compounds, DE-A-2916711, for example, discloses that glycine, α- or β-alanine, proline, glutamine, and the like have a stabilization effect for the blood coagulation factors, and U.S. Pat. No. 4,440,679 describes that arginine, lysine, and/or glycine has a stabilization effect for the VIII factor, fibronectin, and the like. DE-A-1985644 also discloses that arginine, lysine, histidine, phenylalanine, tryptophan, tyrosine, aspartic acid, and/or glutamic acid has a stabilization effect for antithrombin III. However, the stabilization effects of these amino acids are specific to the target proteins or peptides, and therefore, amino acids or combinations of two or more kinds of amino acids having the most effective stabilization effect for arbitrary proteins or peptides cannot be predicted from these findings.
As for FSH, for example, it is described in Patent document 1 that methionine stabilizes gonadotropin in an aqueous composition. Further, as for the stabilization effect of histidine for proteins, it is known that, for example, histidine has a stabilization effect in liquid preparations of blood coagulation factor-related protein (Patent document 2), immunoglobulin (Patent document 3), erythropoietin (Patent document 4), and the like. Furthermore, Patent document 5 discloses a FSH preparation containing glycine, methionine, a nonionic surfactant, and a phosphate buffering agent as stabilizing agents. However, it is not known so far whether histidine has a stabilization effect for FSH in an aqueous composition. It is also not known whether a combination of histidine and another amino acid has a stabilization effect for FSH in an aqueous composition.